Savvas Petanidis

Differential expression of IL-17, 22 and 23 in the progression of colorectal cancer in patients with K-ras mutation: Ras signal inhibition and crosstalk with GM-CSF and IFN-γ.

Recent studies have suggested that aberrant K-ras signaling is responsible for triggering immunological responses and inflammation-driven tumorigenesis. Interleukins IL-17, IL-22, and IL-23 have been reported in various types of malignancies, but the exact mechanistic role of these molecules remains to be elucidated. Given the role of K-ras and the involvement of interleukins in colorectal tumorigenesis, research efforts are reported for the first time, showing that differentially expressed interleukin IL-17, IL-22, and IL-23 levels are associated with K-ras in a stage-specific fashion along colorectal cancer progression. Specifically, a) the effect of K-ras signaling was investigated in the overall expression of interleukins in patients with colorectal cancer and healthy controls, and b) an association was established between mutant K-ras and cytokines GM-CSF and IFN-γ. The results indicate that specific interleukins are differentially expressed in K-ras positive patients and the use of K-ras inhibitor Manumycin A decreases both interleukin levels and apoptosis in Caco-2 cells by inhibiting cell viability. Finally, inflammation-driven GM-CSF and IFN-γ levels are modulated through interleukin expression in tumor patients, with interleukin expression in the intestinal lumen and cancerous tissue mediated by aberrant K-ras signaling. Collectively, the findings a) indicate that interleukin expression is influenced by ras signaling and specific interleukins play an oncogenic promoter role in colorectal cancer, highlighting the molecular link between inflammation and tumorigenesis, and b) accentuate the interwoven molecular correlations as leads to new therapeutic approaches in the future.

Mechanisms and Αpplications of Ιnterleukins in Cancer Immunotherapy

Over the past years, advances in cancer immunotherapy have resulted in innovative and novel approaches in molecular cancer diagnostics and cancer therapeutic procedures. However, due to tumor heterogeneity and inter-tumoral discrepancy in tumor immunity, the clinical benefits are quite restricted. The goal of this review is to evaluate the major cytokines-interleukins involved in cancer immunotherapy and project their basic biochemical and clinical applications. Emphasis will be given to new cytokines in pre-clinical development, and potential directions for future investigation using cytokines. Furthermore, current interleukin-based approaches and clinical trial data from combination cancer immunotherapies will also be discussed. It appears that continuously increasing comprehension of cytokine-induced effects, cancer stemness, immunoediting, immune-surveillance as well as understanding of molecular interactions emerging in the tumor microenvironment and involving microRNAs, autophagy, epithelial-mesenchymal transition (EMT), inflammation, and DNA methylation processes may hold much promise in improving anti-tumor immunity. To this end, the emerging in-depth knowledge supports further studies on optimal synergistic combinations and additional adjuvant therapies to realize the full potential of cytokines as immunotherapeutic agents.

MiR-205 and miR-218 expression is associated with carboplatin chemoresistance and regulation of apoptosis via Mcl-1 and Survivin in lung cancer cells.

Lung cancer chemoresistance is the most frequent barrier in lung cancer therapy. Recent studies have indicated that microRNAs play a significant role in this mechanism and can function as either tumor suppressor or tumor promoters. However the effect of miRNA in lung cancer chemoresistance is poorly understood. Therefore, in the present study we investigated the role of two distinct miR members, the miR-205 and the tumor suppressor miR-218 in the proliferation, invasion and induction of apoptosis in lung cancer cells after carboplatin treatment. The results showed that miR-205 overexpression in A549 and H1975 lung cancer cells is concurrent with the down regulation of miR-218 and in linked with carboplatin sensitivity and chemoresistance. Interestingly, ectopic miR-218 overexpression reduced cell proliferation, invasion and migration of lung cancer cells, whereas miR-205 rescued the suppressive effect of miR-218 by altering the expression levels of the pro-apoptotic proteins PARP, Caspase 3, Bax and upregulating the anti-apoptotic markers Mcl-1 and Survivin. Taken together our findings imply that the miRNAs miR-205 and miR-218 play a key role in the development of lung cancer acquired chemoresistance and the tumor suppressor role of miR-218 in inhibiting lung cancer cell tumorigenesis and overcoming platinum chemoresistance is significant for future cancer therapeutic approaches.

In vitro and ex vivo vanadium antitumor activity in (TGF-β)-induced EMT. Synergistic activity with carboplatin and correlation with tumor metastasis in cancer patients

Epithelial to mesenchymal transition (EMT) plays a key role in tumor progression and metastasis as a crucial event for cancer cells to trigger the metastatic niche. Transforming growth factor-β (TGF-β) has been shown to play an important role as an EMT inducer in various stages of carcinogenesis. Previous reports had shown that antitumor vanadium inhibits the metastatic potential of tumor cells by reducing MMP-2 expression and inducing ROS-dependent apoptosis. However, the role of vanadium in (TGF-β)-induced EMT remains unclear. In the present study, we report for the first time on the inhibitory effects of vanadium on (TGF-β)-mediated EMT followed by down-regulation of ex vivo cancer stem cell markers. The results demonstrate blockage of (TGF-β)-mediated EMT by vanadium and reduction in the mitochondrial potential of tumor cells linked to EMT and cancer metabolism. Furthermore, combination of vanadium and carboplatin (a) resulted in synergistic antitumor activity in ex vivo cell cultures, and (b) prompted G0/G1 cell cycle arrest and sensitization of tumor cells to carboplatin-induced apoptosis. Overall, the findings highlight the multifaceted antitumor action of vanadium and its synergistic antitumor efficacy with current chemotherapy drugs, knowledge that could be valuable for targeting cancer cell metabolism and cancer stem cell-mediated metastasis in aggressive chemoresistant tumors.

Cadmium modulates H-ras expression and caspase-3 apoptotic cell death in breast cancer epithelial MCF-7 cells

Cadmium (Cd) is a well-known metal carcinogen associated with tumor formation and carcinogenesis. It has been shown to induce cancer through various cellular mechanisms involving inhibition of DNA repair, abnormal gene expression, induction of oxidative stress, and triggering apoptosis. It is well-established that the H-ras oncogene is involved in the process of carcinogenesis with direct effects on cellular proliferation and tumorigenesis. Given the biotoxicity of cadmium and its association with carcinogenesis, the effect of that metal ion (Cd(II)) was investigated, in a concentration-dependent fashion, on cell viability, cell proliferation, caspase-3 mediated apoptosis and H-ras gene expression in human breast cancer epithelial MCF-7 cells transfected with the H-ras oncogene (wild type and G12V mutation). The findings show a significant modulation effect of cadmium on H-ras gene expression accompanied by up-regulation of caspase-3-related apoptosis in the concentration range of 100-1000 nΜ cadmium. Concurrently, there is a decrease in MCF-7 proliferation. Collectively, the results a) indicate an interplay of cadmium with H-ras(wt and G12V), with cadmium exhibiting a significant concentration-dependent effect on the modulation of H-ras expression, cell viability and proliferation, and b) project distinctly interwoven roles for both cadmium and H-ras in aberrant physiologies in cancer cells.

Novel ternary vanadium-betaine-peroxido species suppresses H-ras and matrix metalloproteinase-2 expression by increasing reactive oxygen species-mediated apoptosis in cancer cells

Vanadium is known for its antitumorigenicity. Poised to investigate the impact of well-defined forms of vanadium on processes and specific biomolecules (oncogenes-proteins) involved in cancer cell physiology, a novel ternary V(V)-peroxido-betaine compound was employed in experiments targeting cell viability, apoptosis, reactive oxygen species (ROS) production, H-ras signaling, and matrix metalloproteinase-2 (MMP-2) expression in human breast cancer epithelial and lung adenocarcinoma cells. The results reveal that vanadium imparts a significant decrease in cancer cell viability, reducing H-ras and MMP-2 expression by increasing ROS-mediated apoptosis, distinctly emphasizing the nature, structure and properties of ternary ligands on vanadium anti-tumor activity and its future potential as a metallodrug.

Role of Vanadium in Cellular and Molecular Immunology: Association with Immune-Related Inflammation and Pharmacotoxicology Mechanisms.

Over the last decade, a diverse spectrum of vanadium compounds has arisen as anti-inflammatory therapeutic metallodrugs targeting various diseases. Recent studies have demonstrated that select well-defined vanadium species are involved in many immune-driven molecular mechanisms that regulate and influence immune responses. In addition, advances in cell immunotherapy have relied on the use of metallodrugs to create a “safe,” highly regulated, environment for optimal control of immune response. Emerging findings include optimal regulation of B/T cell signaling and expression of immune suppressive or anti-inflammatory cytokines, critical for immune cell effector functions. Furthermore, in-depth perusals have explored NF-κB and Toll-like receptor signaling mechanisms in order to enhance adaptive immune responses and promote recruitment or conversion of inflammatory cells to immunodeficient tissues. Consequently, well-defined vanadium metallodrugs, poised to access and resensitize the immune microenvironment, interact with various biomolecular targets, such as B cells, T cells, interleukin markers, and transcription factors, thereby influencing and affecting immune signaling. A synthetically formulated and structure-based (bio)chemical reactivity account of vanadoforms emerges as a plausible strategy for designing drugs characterized by selectivity and specificity, with respect to the cellular molecular targets intimately linked to immune responses, thereby giving rise to a challenging field linked to the development of immune system vanadodrugs.

In vitro neurotoxic Fe(III) and Fe(III)-chelator activities in rat hippocampal cultures. From neurotoxicity to neuroprotection prospects

It is well known that iron dysregulation is involved in a number of processes involving genetic and non-genetic factors leading to neurodegeneration. Molecules bearing iron or influencing iron metabolism reflect directly into the levels of that redox active metal, present as Fe(II)/Fe(III), in the brain. In turn, iron level variations are associated with chemical reactivity disrupting iron homeostasis, generating variable neurotoxic iron forms and contributing to the vulnerability of cells toward oxidative stress and neuronal death in Alzheimers disease (AD). Efforts to delineate the interactions of neurotoxic Fe(III) with low molecular mass substrates, relevant to cellular processes, led to the discovery of specific well-defined binary iron-quinate (FeQ) species. Poised to investigate the specific effects of a) well-defined forms of labile soluble Fe(III), b) the nature and chemistry of the ligand bound to Fe(III), and c) a natural metal ion binder - quinic acid - acting as a potential neuroprotectant toward iron toxicity, FeCl(3), FeQ, and free quinate were employed in in vitro studies involving primary rat hippocampal cultures. Three hour and 24-hour exposures of such cultures to Fe(III) reveal significant differential effects on both glial and neuronal cell survival linked to neurotoxicity of the specific yet variably composed complex forms of iron. The use of quinic acid both in the free and bound form to Fe(III) a) exemplifies essential structural and chemical attributes of naturally encountered metal ion binders promoting well-defined interactions with neurotoxic Fe(III), and b) signifies the potential linkage of labile Fe(III) chemical reactivity in neurodegeneration with natural substrate neuroprotection.

Potential synergistic effect of phosphodiesterase inhibitors with chemotherapy in lung cancer

Purpose: Lung cancer remains the leading cause of cancer-related deaths worldwide and novel therapeutic approaches targeting crucial pathways are urgently needed to improve its treatment. Differentiation-based therapeutics (Methylxanthines) and phosphodiesterase inhibitors (type 4 and 5), have been implicated in cancer treatment. Our objectives were to capture any potential anti-tumor effect of these drug combinations with chemotherapeutic agents in vitro. Methods: Theophylline as Methylxanthines, Roflumilast as phosphodiesterase type 4 (PDE4) inhibitor and Sildenafil as phosphodiesterase type 5 (PDE5) inhibitor are the drugs that we combined with the chemotherapeutic agents (Docetaxel, Cisplatin and Carboplatin) in vitro. Lung cancer cell lines (NCI-H1048-Small cell lung cancer-SCLC, A549- Non-small cell lung cancer-NSCLC) were purchased from ATCC LGC Standards. At indicated time-point, following 24h and 48h incubation, cell viability and apoptosis were measured with Annexin V staining by flow cytometry. Statistical analysis was performed by GraphPad Prism. Results: In SCLC, following 48h incubation, platinum combinations of carboplatin with roflumilast and sildenafil (p<0.001) and carboplatin with theophylline and sildenafil showed increased apoptosis when compared to carboplatin alone. Concerning the combinations of cisplatin, when combined with roflumilast, theophylline and sildenafil appeared with increased apoptosis of that alone (p<0.001, 24h and 48h incubation). In NSCLC, the 24h incubation was not enough to induce satisfactory apoptosis, except for the combination of cisplatin with roflumilast and theophylline (p<0.05) when compared to cisplatin alone. However, following 48h incubation, carboplatin plus sildenafil, carboplatin plus sildenafil, theophylline and roflumilast showed more cytotoxicity when compared to carboplatin alone (p<0.001). Docetaxel combinations showed no statistically significant results. Conclusion: The synergistic effect of PDE inhibitors with platinum-based agents has been demonstrated in lung cancer. Our suggestion is that these combinations could be used as additive and maintenance treatment in combination to antineoplastic agents in lung cancer patients.

Autophagy inhibition upregulates CD4+ tumor infiltrating lymphocyte expression via miR‐155 regulation and TRAIL activation

Chemoresistance is a major challenge in lung cancer treatment. Recent findings have revealed that autophagic mechanism contributes significantly to immunosuppressive related chemoresistance. For that reason, targeting autophagy‐related immunosuppression is an important approach to reverse tumor drug resistance. In this study, we report for the first time that autophagy inhibition triggers upregulation of CD4+, Foxp3+ tumor infiltrating lymphocytes in late metastatic lung cancer tissues. Furthermore, autophagy blockage induces chemosensitization to carboplatin, immune activation and cell cycle arrest. This induction correlated with reduction in expression of drug resistance genes MDR1, MRP1, ABCG2 and ABCC2 along with decreased expression of PD‐L1 which is associated with severe dysfunction of tumor specific CD8+ T cells. Furthermore, experiments revealed that co‐treatment of carboplatin and autophagy inhibitor chloroquine increased lung tissue infiltration by CD4+, FoxP3+ lymphocytes and antigen‐specific immune activation. Subsequent ex vivo experiments showed the activation of carboplatin related TRAIL‐dependent apoptosis through caspase 8 and a synergistic role of miR‐155 in lung tissue infiltration by CD4+, and FoxP3+ lymphocytes. Overall, our results indicate that autophagy blockage increases lung cancer chemosensitivity to carboplatin, but also reveal that miR‐155 functions as a novel immune system activator by promoting TILs infiltration. These results indicate that targeting of autophagy can prevent cancer related immunosuppression and elucidate immune cell infiltration in tumor microenvironment thus representing a potential therapeutic strategy to inhibit lung cancer progression and metastasis.